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2004 Hindlimb function in the alligator: integrating movements, motor patterns, ground reaction forces and bone strain of terrestrial locomotion Stephen M. Reilly

Jeffrey S. Wiley

Audrone R. Biknevicius

Richard W. Blob Clemson University, [email protected]

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This Article is brought to you for free and open access by the Biological Sciences at TigerPrints. It has been accepted for inclusion in Publications by an authorized administrator of TigerPrints. For more information, please contact [email protected]. The Journal of Experimental Biology 208, 993-1009 993 Published by The Company of Biologists 2005 doi:10.1242/jeb.01473

Hindlimb function in the alligator: integrating movements, motor patterns, ground reaction forces and bone strain of terrestrial locomotion Stephen M. Reilly1,*, Jeffrey S. Willey1, Audrone R. Biknevicius3 and Richard W. Blob2 1Department of Biological Sciences, Ohio University, Athens, OH 45701, USA, 2Department of Biological Sciences, Clemson University, Clemson, SC 29634, USA and 3Department of Biomedical Sciences, Ohio University College of Osteopathic Medicine, Athens, OH 45701, USA *Author for correspondence (e-mail: [email protected])

Accepted 21 December 2004

Summary Alligator hindlimbs show high torsional loads during during stance phase in sprawling and erect quadrupeds terrestrial locomotion, in sharp contrast to the bending or act in isometric or even eccentric contraction in alligators, axial compressive loads that predominate in animals that stabilizing the knee and ankle during the support-and- use parasagittal limb movements. The present study propulsion phase. Motor patterns in alligators reveal the integrates new data on hindlimb muscle function with presence of local and temporal segregation of muscle previously obtained data on hindlimb kinematics, motor functions during locomotion with muscles that lie side by patterns, ground reaction forces and bone strain in order side dedicated to performing different functions and only to (1) assess mechanisms underlying limb bone torsion one of 16 muscles showing clear bursts of activity during during non-parasagittal locomotion in alligators and (2) both stance and swing phases. Data from alligators add to improve understanding of hindlimb dynamics during other recent discoveries that homologous muscles across terrestrial locomotion. Three dynamic stance phase quadrupeds often do not move joints the same way as is periods were recognized: limb-loading, support-and- commonly assumed. Although alligators are commonly propulsion, and limb-unloading phases. Shear stresses due considered models for early semi-erect tetrapod to torsion were maximized during the limb-loading phase, locomotion, many aspects of hindlimb kinematics, muscle during which the ground reaction force (GRF) and activity patterns, and femoral loading patterns in caudofemoralis (CFL) muscles generated opposing alligators appear to be derived in alligators rather than moments about the femur. Hindlimb retraction during reflecting an ancestral semi-erect condition. the subsequent stance-and-propulsion phase involves substantial medial rotation of the femur, powered largely by coordinated action of the GRF and CFL. Several Key words: locomotion, kinematics, kinetics, bone strain, motor muscles that actively shorten to flex and extend limb joints patterns, alligator.

Introduction Terrestrial tetrapods use diverse limb postures during broad patterns in the mechanics and dynamics of tetrapod limb locomotion, ranging from the highly sprawling posture of movements (e.g. Cavagna et al., 1977; Biewener 1989, 1990; salamanders and some lizards (in which the limbs are held out Alexander and Jayes, 1983; Hildebrand, 1976; Demes et al., to the side of the body; Ashley-Ross, 1994a,b; Reilly and 1994; Blickhan and Full, 1992). However, an increasing DeLancey; 1997a,b; Rewcastle, 1981) to the parasagittal number of studies on limb function in tetrapods using non- posture of birds and many mammals (in which the limbs are parasagittal limb postures have identified major differences in held more directly beneath the body; Jenkins, 1971; Reilly, limb mechanics between animals that use parasagittal limb 2000; Gatesy, 1990). Between these extremes, several species movements versus those that use non-parasagittal limb use intermediate postures (Jenkins, 1971; Gatesy, 1997; movements (Blob and Biewener, 1999, 2001; Reilly and Blob, Pridmore, 1985), and some, such as crocodilians and some 2003; Parchman et al., 2003; Willey et al., 2004). Among the lizards (e.g. iguanas), are capable of using a range of limb foremost of these differences is the contrasting loading regimes postures even over a restricted range of speeds (Gatesy, 1991; found in the limb bones. Although there are some exceptions Reilly and Elias, 1998; Blob and Biewener, 1999, 2001; Reilly (e.g. the femora of chickens; Carrano, 1998), the limb bones and Blob, 2003). Studies of several different aspects of of species using parasagittal limb movements are primarily terrestrial locomotion in mammalian and avian species using loaded in bending or axial compression (Alexander, 1974; parasagittal limb postures provided a foundation for discerning Rubin and Lanyon, 1982; Biewener et al., 1983, 1988). In

THE JOURNAL OF EXPERIMENTAL BIOLOGY 994 S. M. Reilly and others contrast, torsional loads exceed bending loads in species using bipedal species (kangaroos, birds, humans). To date, no study non-parasagittal posture that have been examined to date has combined analyses of locomotor forces, limb bone loads, (alligators and iguanas; Blob and Biewener, 1999, 2001). motor patterns, kinematics and foot fall patterns to evaluate Though torsional loads are clearly present in the limb bones how a non-erect quadrupedal animal supports its body weight of the non-parasagittal species that have been tested, the and generates propulsive forces with its limbs. underlying causes of this torsion remain to be clarified. In the In this study, we present an integrative analysis of few vertebrates that have been found to exhibit strong torsional propulsive dynamics in the hindlimb of the American alligator loading of limb bones (e.g. walking chickens, flying bats and (Alligator mississippiensis Daudin). By integrating data on birds; Pennycuick, 1967; Swartz et al., 1992; Biewener and limb forces, bone loads, kinematics and motor patterns, we are Dial, 1995; Carrano, 1998), these torsional loads are believed able to evaluate specific questions about locomotor mechanics to be induced by locomotor forces acting at a distance from the in this species. In particular, we assess the potential long axis of the limb bone which, therefore, generate a mechanisms underlying limb bone torsion during non- torsional moment. In species using non-parasagittal terrestrial parasagittal locomotion in alligators. Additionally, our locomotion, two different forces are likely to act at a distance analyses allow us to refine evaluations of limb muscle function from limb bone long axes and potentially contribute to and the roles that these muscles play in supporting body weight torsional moments. First, because species using non- and retracting the limb. Based on limb movement, force and parasagittal locomotion hold their limbs out to the side of the motor pattern data, we distinguish three dynamic phases during body for much of limb support, the ground reaction force hindlimb stance: a limb-loading phase, a support-and- (GRF) will be directed either anterior or posterior to the long propulsive phase, and a limb-unloading phase. Comparisons of axis of limb bones for most of the step in these animals (Blob locomotor patterns among tetrapods suggest that limb function, and Biewener, 1999, 2001). At the same time, the GRF will be forces, and the use of femoral rotation in alligators may be directed more nearly perpendicular to the femur of non- quite different from patterns understood for other species that parasagittal animals than it will be in parasagittal animals. As use either more sprawling or more erect limb posture. These a result, the GRF will tend to rotate limb bones about their long results suggest that species using non-parasagittal posture may axis in these species, contributing to torsional loading. Second, exhibit considerable diversity in their locomotor mechanics. the major limb retractor muscle in many non-parasagittal Furthermore, tail dragging has been shown to have serious species (including alligators and iguanas) is the caudofemoralis consequences for locomotor mechanics in alligators (Willey et longus (CFL), which inserts on the ventral surface of the al., 2004) which are borne out in patterns of femoral function proximal femur (Snyder, 1962; Reilly, 1995; Gatesy, 1997). in this study. Thus, distinctive features of locomotor dynamics When this muscle contracts to retract the leg during the stance in alligators may be a consequence of dragging the tail rather phase of locomotion, it acts at a distance from the long axis of than general features of non-parasagittal postures. the femur equal to the radius of the bone, and this distance is a moment arm for long axis rotation of the femur. Thus, contraction of CFL should produce inward (medial) rotation of Materials and methods the femur and torsional loading. This analysis re-examines and synthesizes data that were Though both mechanisms for the induction of torsion are collected during the course of several studies of terrestrial viable, the relative contributions of each (and their potential locomotion in alligators, including studies of kinematics and interactions) are uncertain. As a result, it is difficult to predict gait (Reilly and Elias, 1998), in vivo limb bone strains (Blob whether torsion should be expected to predominate in the limb and Biewener, 1999), locomotor forces (Blob and Biewener, bones of all species that use non-parasagittal limb movements 2001; Willey et al., 2004), and hindlimb muscle activity or whether specific differences in limb movements or patterns (Gatesy, 1997; Reilly and Blob, 2003). Details of the mechanics might lead to alternative expectations for some methods used for data collection and initial analyses can be lineages. To evaluate the contributions of these mechanisms to found in those publications. In the sections that follow, we torsional loading of limb bones during locomotion, several summarize information on the methods used in these studies types of data must be integrated, including information on limb that are most relevant to our present analysis. Animals used in position and movements, locomotor forces, muscular action all of the studies on which our present analyses are based were and bone loading. The integration of such diverse sets of data provided by the Rockefeller Wildlife Refuge (Grand Chenier, is crucial not only for a complete understanding of bone LA, USA). All experimental procedures complied with loading mechanics, but also, ultimately, for a thorough IACUC guidelines for the institutions where data were understanding of the functional dynamics of how animals collected. control forward propulsion with their limbs. Data on locomotor In our synthesis of data from different sources, we based our dynamics are available for only a narrow range of vertebrate analyses on trials that were collected from animals as similar taxa. Though studies focusing on multiple levels of analysis in size as possible, under locomotor situations that were as have provided major insights into propulsive dynamics in similar as possible. In general, data were collected at tetrapods, most studies have focused on animals with erect temperatures between 22°C and 29°C, from subadult postures (cursorial mammals, primates) and, in particular, individuals 0.5–1.0·m in length and 0.5–4.0·kg in mass, at

THE JOURNAL OF EXPERIMENTAL BIOLOGY Alligator hindlimb function 995 speeds from 0.1–0.6·m·s–1, using a standard ‘high walk’ limb speed variation among strides for all individuals was posture (limbs adducted 55±5° below horizontal at mid- 0.141–0.151·m·s–1. This is less than 7% variation among stance), and a duty factor (ratio of hindlimb stance duration to strides, well within the range of variation reported in previous stride duration) of approximately 0.7. Specifics for animal studies of alligator kinematics (Gatesy, 1991) and muscle sizes, locomotor speeds and gait parameters are noted in the activity patterns (Gatesy, 1997). Electromyographical data descriptions of methods provided for each set of experimental were collected from these exact same strides. data. Although there is variation in some of these parameters, Reflective landmarks (2·mm diameter dots visible in both many were very closely matched across studies (e.g. mean the lateral and dorsal views: Fig.·1A,B) were painted on the duty factors across kinematic, force, bone strain and skin of the alligators to mark positions along the vertebral electromyography datasets ranged from 0.70–0.73). Moreover, column (T), the hip joints (directly over the acetabula; H), and we believe that over the extensive ranges of size and behavior three landmarks on the right hindlimb: the knee joint (on the of alligators, variation among the individuals and trials in our anterolateral point of the knee when flexed; K), the ankle joint analyses is minimal, and that the insights gained from our (posterolateral point of the ankle when flexed; A) and the foot synthesis outweigh the potential complications that arise from (lateral aspect of the metatarsal– tarsal articulation; F). Three- the combination of data collected during different experiments. dimensional coordinates of each landmark were digitized using stereo measurement TV (sMTV; Updegraff, 1990), and Kinematics and gait analyses kinematic angles were calculated from these coordinates with Kinematic angles and gait patterns were calculated and an accuracy of ±1° for each joint. graphed to indicate the general position and direction of The knee and ankle angles calculated were the actual three- movement of the limb segments. Data on hindlimb kinematics dimensional angles for these joints based on the landmarks and gait patterns were collected from five alligators (total above (Fig.·1B,C). Femoral movements were quantified length 0.48–0.54·m, body mass 247–333·g); detailed analyses using two three-dimensional angles: femoral retraction for the strides used in the present analysis are presented in (retraction/protraction movements relative to the longitudinal Reilly and Elias (1998). Alligators were filmed under strobe axis of the pelvis) and femoral adduction (adduction/abduction lights at 200·fields·s–1 using a NAC HSV-400 high-speed video position relative to the mediolateral axis of the pelvis). Femoral system. Both lateral and dorsal views of the alligators were retraction was measured as the angle between the femur and filmed (using mirrors) as they ‘high walked’ on a 70·cm long a line from the acetabulum to the trunk landmark. This canvas treadmill. Only strides during which the animals very calculation produces angles that are 5–10° greater (not 15° as nearly matched the speed of the treadmill (0.146·m·s–1) were indicated by Reilly and Elias, 1998) than those that would be analyzed, during which the position of a landmark painted on calculated if femoral position were measured relative to the the hip stayed within a 1·cm zone (i.e. ±0.005·m). Based on sagittal plane (e.g. Gatesy, 1991) but produces kinematic the measured durations of these strides, the complete range of profiles that are nearly identical (within 5–10%; Reilly and

0.6 A D Peak vertical force H T 0.5 H K 0.4 Vertical A F Vertebral axis 0.3 H (left) Peak braking Pelvis Pelvis 0.2 force Peak Knee H Transverse axis Braking propulsive H H to propulsive force angle through 0.1 transition T acetabula (Hip left & right) 0 Fore–aft Hip K Hip 0255075100 A retraction Hind limb forces (BWU) –0.1 Lateral angle adduction K F Ankle Peak lateral force angle –0.2 B angle C Stance duration

Fig.·1. Kinematic landmarks and angles used to describe hindlimb movements in alligators during locomotion. (A) Three-dimensional coordinates were digitized for one trunk landmark (T), and landmarks for the hip joint (H; on both sides: H left, H right); the knee (K), the ankle (A) and foot (F; on the skin on the lateral aspect of the metatarsal–tarsal articulation). (B) Three-dimensional angles were calculated for femoral retraction angle (the angle between landmarks T–H right-K indicating primarily femoral retraction/protraction movements); knee angle (angle H–K–A indicating knee flexion and extension), and ankle angle (angle K–A–F indicating foot flexion and extension). (C) Femoral adduction angle (angle H left-H right-K) was calculated to quantify the degree of hip adduction relative to a transverse line through the acetabula. (D) Ground reaction forces in body weight units (BWU) of a typical trial indicating key kinetic events: peak vertical force, peak braking force, braking-to-propulsive transition (zero fore–aft force), peak propulsive force and peak lateral force.

THE JOURNAL OF EXPERIMENTAL BIOLOGY 996 S. M. Reilly and others

Elias, 1998). Femoral adduction was measured as the angle metal tips of each bifiler insulated electrode were 0.5·mm long. between the femur and a transverse axis through the acetabula Electrodes were implanted percutaneously through the skin (based on three-dimensional coordinates of both hips), with 0° directly into the belly of each target muscle. The bundle of indicating no femoral adduction (the femur held straight out electrodes was glued together and sutured to a scale on the laterally from the acetabulum) and 90° indicating a position midline dorsal to the pelvis. Animals completely recovered parallel to the sagittal plane of the alligator (Reilly and Elias, from anesthesia within 2·h and all synchronized EMG and 1998). This convention for reporting femoral adduction angles kinematic data were recorded during the next 2·h. Animals follows the evolutionary sprawling-to-erect paradigm, which were rested (about 15–30·min) between bouts of walking (45·s categorizes sprawling femoral angles as 0° and erect ones as maximum). Immediately following each experiment, the test 90° (Bakker, 1971; Charig, 1972; Parrish, 1986, 1987; Reilly animal was sacrificed by overdose of anesthetic and preserved and Elias, 1998; Blob, 2001). Our femoral adduction in 10% formalin. Electrode position was then confirmed by calculations also account for pelvic roll about a longitudinal dissection, and EMG data were considered valid for analysis axis (<6° to each side; Gatesy, 1991) and, thus, effectively only for preparations in which the electrode lay completely represent the angle between the femur and the horizontal plane within the muscle. of the body of the alligator. However, from a practical EMG signals were amplified 10·000 times using AM perspective, because pelvic roll is not very large in alligators Systems model 1700 differential AC amplifiers with a (Gatesy, 1991) the difference between the convention of Reilly bandpass of 100–3000·Hz (and a 60·Hz notch filter) and then and Elias (1998) that we use in this analysis and conventions recorded on a TEAC XR-5000 multichannel FM tape recorder that refer to absolute planes is minimal. along with a synchronization pulse simultaneously recorded on Gaits used by alligators were identified by limb phase the video frames. The analog signals (EMG channels plus a (Hildebrand, 1976; Reilly and Biknevicius, 2003), the elapsed synchronization pulse) for each stride were converted to a time between a hindlimb strike and the ipsilateral forelimb digital data file using custom software with a Keithley analog- strike normalized to hindlimb stride duration. Gaits used to-digital converter and a microcomputer. The effective sample during kinematic, electromyographic, force platform and bone rate for each channel was 10·kHz at 12-bit resolution. A strain experiments had nearly identical duty factors and limb 10·kHz sample rate was used because previous work has phase relationships. shown that this rate allows the faithful reproduction of EMG For the analyses performed in this study, mean kinematic spikes from vertebrate locomotor muscles (Jayne et al., 1990). profiles and gait patterns for three-dimensional joint Prior to the experiments, an extensive calibration of the system movements during high walks were taken from figs·4 and 5 of revealed no crosstalk downstream of the electrodes, and Reilly and Elias (1998). These plots were aligned with data on crosstalk has not been a problem in previous work using limb forces, limb bone strains, and limb muscle activity in the same electrode materials, construction and placement order to evaluate how limb forces and their bone loading protocols. EMG profiles were inspected for possible patterns consequences are produced. revealing crosstalk, and none were found. Ten electrodes were implanted during each EMG Electromyography experiment and, of these, five to six usually supplied successful To quantify patterns of activity (motor patterns) for alligator data recordings, providing data from between one and three hindlimb muscles during the high walk, electromyographic individuals for each target muscle (Table·1). Although we did (EMG) recordings were collected from 12 muscles (all on the not record from every target muscle in each of our five right side of the body). A previous study (Reilly and Blob, experimental alligators, we were able to record data from 2003) examined EMG data from the same alligator multiple individuals for seven muscles and it is upon these data experiments but sampled muscle activity patterns across a that we based our primary conclusions. Custom software was range of femoral angles from 19–55° and compared ~30° to used to digitize the times of burst onset and offset for each ~50° patterns to test for statistical effects of posture on the muscle relative to the timing of foot down within each stride. modulation of the timing and amplitudes of motor patterns. This was done to assess changes in muscle activity relative to The present study presents new EMG data for the alligators the onset of stance phase (i.e. when the foot contacts the ground normal high walk posture (femurs adducted 55±5° below and limb movements begin to directly affect the propulsive horizontal at mid-stance), from the exact same strides (10 each dynamics of the limb cycle). To quantitatively characterize from five animals of total length 0.48–0.54·m, body mass patterns of muscle activity, calculations of average burst 247–333·g; speed (0.141-0.151·m·s–1) of treadmill locomotion patterns (EMG bars on Fig.·3F) were based on 10 strides examined in the synchronized kinematic analyses described from each individual for which muscles were successfully above (Reilly and Elias, 1998). implanted. To facilitate the calculation of average EMG EMG recordings were made from bipolar stainless steel patterns among strides, and allow alignment of EMG, electrodes implanted into each muscle as in previous research kinematic and gait data with force and strain datasets, durations (Reilly, 1995). All electrodes were implanted while the animals of muscle bursts were scaled as a percentage of stride duration. were under anesthesia induced by placing the animals in a To gain more complete insight into muscular contributions closed container with 1·ml of halothane for 15·min. Bared to locomotor movements, forces and bone loading patterns

THE JOURNAL OF EXPERIMENTAL BIOLOGY Alligator hindlimb function 997

(including femoral torsion), we supplemented our own A C ILFIB* recordings of 12 muscle activity patterns with EMG data for four additional muscles published by Gatesy (1997). Details of the data collection methods for those muscles (caudofemoralis FTI2* PIFE3 ILFEM longus, flexor tibialis head 2, iliofibularis, and CFL* puboischiofemoralis externus head 2; indicated by asterisks in Table·1 and Figs·2, 3) are provided in Gatesy’s original FTE ADD1 PIT publication. Gatesy’s data (Gatesy, 1997) can be reasonably compared to those for the other muscles examined in this D analysis because Gatesy also collected EMGs during treadmill PIFI2 walking, and he recorded from animals of similar size B ILTIB2 AMB1 (50–70·cm total length) under similar locomotor conditions (speed: 0.1–0.15·m·s–1; duty factor: 0.73±0.03; femoral ILTIB1 adduction angle: 60°) to those used here. Gatesy’s study FMTI PIFE2* (Gatesy, 1997) normalized burst timings to stride duration, but G reported values relative to the onset of the caudofemoralis TA muscle burst. To coordinate data from Gatesy’s study with EMGs from Reilly and Blob (2003) and force and strain records, Gatesy’s burst timing data were adjusted by Fig.·2. Alligator hindlimb muscles involved in the high walking for subtracting 6% from all onset and offset values (adjustment which electromyographical data have been recorded. Lines of actions based on fig.·5 in Gatesy, 1997), thus realigning his EMG are indicated for muscles active in either stance phase (red), swing records relative to the beginning of stance phase. It should also phase (blue) or with activity in both phases (black); gray shading in be noted that eight muscles we recorded were also examined arrows indicates that the muscle lies medial to the femur. AMB1, by Gatesy (1997). For two of these (flexor tibialis externus, ambiens, head 1; ADD1, adductor femoris 1; CFL, caudofemoralis iliotibialis head 2) our data provide positive verification of longus; FMTI, femorotibialis internus; FTE, flexor tibialis externus; weak or sporadic patterns found by Gatesy (1997), and for the FTI2, flexor tibialis internus, head 2; G, gastrocnemius; ILFEM, remaining six muscles EMG patterns were similar in both iliofemoralis; ILFIB, iliofibularis; ILTIB1/2 iliotibialis, head 1/2; studies. PIFE2/3, puboischiofemoralis externus, head 2/3; PIFI2, puboischiofemoralis internus, head 2; PIT, puboischiotibialis; TA, Myology tibialis anterior. Data for muscles marked with asterisks are taken from Gatesy (1997). The continuation of the AMB1 tendon through By combining EMG data from Gatesy (1997) with our EMG the extensor sheet and into the Achilles tendon is not shown. data, we were able to coordinate examination of the motor patterns of 16 alligator hindlimb muscles (including axial, thigh, and crural muscles) with kinematic, gait, force and bone most posteriorly located muscle of the posterior thigh. FTE strain data. Detailed anatomical descriptions of most of these also functions as a knee flexor and ankle extensor muscles (except the tibialis anterior) have been published (plantarflexor). (Romer, 1923; Gatesy, 1997). However, to aid understanding Flexor tibialis internus, head 2 (FTI2; Fig.·2A; two of muscle function and our analyses, in this section we provide individuals). Origin: postacetabular iliac blade. Insertion: a brief summary of muscle morphology (together with a medially on proximal tibia. The FTI2 lies just ventral to FTE schematic illustration, Fig.·2). Previous studies have evaluated and is the largest of the four flexor tibialis internus slips. FTI2 alligator muscle functions based on anatomical topography shares a wide tendon with the puboischiotibialis (PIT) before (Romer, 1923) or EMG patterns (Gatesy, 1997). By correlating inserting with the FTE on the proximal tibia. EMG data from EMGs with limb force and bone strain data, our present Gatesy (1997). analyses allow us to test many of these proposed functions. Caudofemoralis longus (CFL; Fig.·2A; four individuals). Muscles are described in anatomical and functional groups Origin: caudal vertebrae 3–15, filling the space in the tail based on results of Gatesy (1997), Reilly and Blob (2003), and between vertebral haemal arches and transverse processes. this analysis. Refined details of muscle functions found in this Insertion: proximally by a tendon on to the fourth trochanter study are presented in the results. and surrounding area of femur; distally (not illustrated) by a smaller auxiliary tendon to the knee and calf musculature. The Posterodorsal thigh muscles (femoral retractors) CFL is the largest muscle affecting hindlimb movements, Flexor tibialis externus (FTE; Fig.·2A; one individual). producing femoral long axis rotation as well as retraction. Origin: tip of the postacetabular iliac process. Insertion: EMG data from Gatesy (1997). medially by a large tendon to the proximal tibia; distally by a smaller tendon passing down the leg to the calcaneus. It Anterodorsal thigh muscles (knee extensors) extends along the posterior aspect of the femur where it joins Iliotibialis, head 1 (ILTIB1; Fig.·2B; one individual). the heads of the flexor tibialis internus. FTE is the largest and Origin: anterodorsal rim of iliac blade. Insertion: via extensor

THE JOURNAL OF EXPERIMENTAL BIOLOGY 998 S. M. Reilly and others tendon over anterior surface of knee to the proximal tibia. The anterior surface of the proximal thigh. It is larger and originates iliotibialis has three heads originating along the rim of the iliac more anteriorly than PIFI1 (not sampled). blade. All three heads join the distal tendon of femorotibialis Ambiens, head 1 (AMB1; Fig.·2D; two individuals). Origin: internus (FMTI) to form a common extensor tendon inserting ilioischiadic junction anterior to the acetabulum. Insertion: on the front of the tibia. ILTIB1 is the most anterior of the three proximally, over the anterior surface of the knee to the heads, smaller and deep to the ILTIB2. proximal tibia via the common extensor tendon with Iliotibialis, head 2 (ILTIB2; Fig.·2B; two individuals). femorotibialis and iliotibialis; distally, a weaker tendon Origin: anterodorsal rim of iliac blade. Insertion: via extensor continues through the extensor sheet lateral the knee joint, tendon over anterior surface of knee to the proximal tibia. The inserting on the gastrocnemius and Achilles tendon (not ILTIB2 is the middle head of the iliotibialis group. It is the shown). The AMB1 is a large muscle that extends along the largest muscle on the anterodorsal aspect of the thigh and anterior aspect of the thigh. AMB1 is one of two heads of originates from a wide portion of the iliac crest. ambiens that is more superficial and considerably larger than Both the ILTIB1 and 2 may play a role in hip and knee AMB2 (not sampled). stabilization as well as knee extension (Reilly and Blob, 2003). Puboischiofemoralis externus, head 2 (PIFE2; Fig.·2D; two Femorotibialis internus (FMTI; Fig.·2B; three individuals). individuals). Origin: ventral surface of the pubis and last Origin: dorsal surface of central femoral shaft. Insertion: via abdominal rib. Insertion: posteroventral surface of the extensor tendon over anterior surface of knee to the proximal proximal femur. The PIFE2 runs from the anterior to the tibia. The FMTI is the larger, more anterior belly of the two posteroventral aspect of the thigh. It is one of three heads of heads of the femorotibialis group. It shares a broad, tendinous the PIFE group originating from the ventral pelvic elements insertion with femorotibialis externus, iliotibialis and ambiens. that converge to insert on proximal femur. It is located anteriorly to PIFE3 (described above under femoral adductors). Ventral thigh muscles (femoral adductors) EMG data from Gatesy (1997). Adductor femoris 1 (ADD1; Fig.·2C; three individuals). Origin: ventral aspect of ischium. Insertion: ventral shaft of Crural muscles femur. The ADD1 is one of two heads of the adductor femoris Gastrocnemius (G; Fig.·2D; three individuals). Origin: muscle and the most superficial ventral muscle of the thigh. ventral aspect of distal femur and posterior proximal tibia. Puboischiotibialis (PIT; Fig.·2C; 1 individual). Origin: Insertion: calcaneal tuber via Achilles tendon. This ankle anterior edge of ischium, well below acetabulum. Insertion: extensor is the largest muscle of the crus and appears to act as medial aspect of tibia. The PIT is a large muscle lying just a knee stabilizer or flexor as well as ankle extensor (Blob and posterior to ADD1. It passes along the ventral surface of the Biewener, 2001; Reilly and Blob, 2003). thigh to join FTI2 before inserting on the tibia. Tibialis anterior (TA; Fig.·2D; two individuals). Origin: Puboischiofemoralis externus, head 3 (PIFE3; Fig.·2C; two Anterior aspect of distal femur and proximal tibia and fibula. individuals). Origin: ventral surface of ischium. Insertion: Insertion: Distal dorsolateral surfaces of the four most medial Posteroventral surface of proximal femur. The three heads of metatarsals. The TA passes through a laterally offset groove in the puboischiofemoralis externus complex converge to a the proximal end of the tibia to act as a dorsiflexor of the ankle. common insertion on the proximal femur but are distinguished by their origins. The first and second heads (PIFE1 and 2) are Hindlimb ground reaction forces more anterior and originate from the pubis and ribs, whereas To evaluate hindlimb ground reaction forces (GRFs) in the PIFE3 is located more posteriorly. context of other datasets, a subset of GRF data reported by Willey et al. (2004) were selected that had been collected under Dorsal thigh muscles (femoral abductors) locomotor conditions as close as possible to those of the Iliofibularis (ILFIB; Fig.·2C; three individuals). Origin: kinematic, gait, and EMG data. GRFs were collected from five central iliac blade. Insertion: Fibular tubercle and alligators (00.99–1.09·m total length; 2.24–4.00·kg body mass) gastrocnemius. The ILFIB extends across the hip and knee as they walked over a force platform (Kistler Corporation, joints parallel to the femur. EMG data from Gatesy (1997). plate Type 9281B) inserted into a 6.1·m runway. To capture Iliofemoralis (ILFEM; Fig.·2C; one individual). Origin: records for individual footfalls of the hindlimb and reduce the postacetabular iliac blade. Insertion: posterodorsal aspect of potential for interference from contacts with other feet, a proximal femur. The ILFEM runs parallel to insert between the triangular insert (244.25·cm) was firmly affixed to the surface origins of the two heads of the femorotibialis group. of the force platform and made flush with the floor of the trackway, with the uninstrumented part of the track covering Anterior thigh muscles (femoral protractors) the rest of the platform. Analog signals from the force platform Puboischiofemoralis internus, head 2 (PIFI2; Fig.·2D; one were amplified (Kistler Corporation, amplifier Type 9865C), individual). Origin: centra of lumbar vertebrae and the ventral digitized at 500·Hz, and imported into Bioware 2.0 software surfaces of their transverse processes. Insertion: Anterodorsal for analysis. Trials were synchronously recorded by two 60·Hz aspect of proximal femur. One of two heads in the cameras (JVC TK-C1380U) oriented to provide views that puboischiofemoralis internus group, PIFI2 lies along the would allow evaluation of whether footfalls were acceptable

THE JOURNAL OF EXPERIMENTAL BIOLOGY Alligator hindlimb function 999 for analysis (e.g. whether a single foot landed fully on the strains (tension or compression along the axis of the bone). The platform without overlapping feet). A single high-speed use of rosette gauges also allowed calculation of the magnitude camera (Redlake Motionscope 500) simultaneously recorded and orientation of principal strains (maximum and minimum the trials from a lateral view at 250·Hz in order to determine strains at a site, potentially not aligned with the long axis of the velocity of each trial. Mean locomotor speed of each trial the bone; Daley and Riley, 1978), as well as shear strains was determined from the time it took the animal to walk past (calculated following methods of Biewener and Dial, 1995). a 70·cm calibration grid located on the back wall of the runway. These calculations allowed evaluation of the importance of Forward speeds were also calculated between successive torsional loading on the femur and the timing of its 10·cm vertical bars immediately over the force platform. These development through the stride cycle. velocities were compared with the mean velocity, and trials After gauge attachment, lead wires from the gauges were were accepted only if the successive velocities differed by less passed subcutaneously out of an incision dorsal to the than 5% from the mean velocity (indicating the animals were acetabulum and soldered into a microconnector for connection not accelerating or decelerating). Duty factor (percentage of to Vishay conditioning bridge amplifiers (model 2120, stride duration that the individual limb contacted the force Measurements Group). After 2–4·days of recovery from platform) also was calculated from force and video records. surgery, data were collected during 40·s bouts of treadmill GRFs were collected in the vertical, fore–aft (craniocaudal), locomotion at either 0.17 or 0.37·m·s–1 [note that Reilly and and mediolateral directions relative to the alligators (Fig.·1D). Elias (1998) found no speed effects on femoral retraction and Vertical forces reflect body support and vertical acceleration adduction kinematics, thus we believe that strain data captured of the center of mass due to gravity. Fore–aft forces are divided at slightly higher speeds should be comparable to the other data into braking (–) or propulsive (+) components. Mediolateral sets]. Raw strain signals were sampled through an A/D forces indicate whether a limb is pushing medially (+) or converter at 100·Hz and stored on computer for analysis. laterally (–). All GRFs were converted to body weight units Locomotor trials were filmed with 60·Hz video to allow (BWU) to adjust for differences in body mass among the evaluation of limb movements, speed, and duty factor for each animals. Five parameters of the GRF and their timing stride. To evaluate the timing of peak shear strains and the (percentage stance duration) were measured: peak vertical development of other strains relative to muscle activity and force, peak braking force, braking-to-propulsive transition limb forces, representative strain traces were normalized for (fore–aft force crosses zero), peak propulsive force, and peak stride duration and aligned with traces for other locomotor lateral force. parameters (see Fig.·4). From the 60 trials (12 runs per animal) analyzed by Willey In addition to strain recordings, force platform records (Blob et al. (2004), 20 high walk trials were selected from four of the and Biewener, 2001) also provided data on femoral torsion by alligators to produce a sample that very closely matched the allowing calculation of the torsional moment of the GRF about mean speed (0.146·m s–1) and duty factor (0.70±0.01) of the the long axis of the femur. These data were obtained from six trials used in the analyses of kinematics, gait, and EMGs. This trials for a single alligator (1.98·kg) moving faster than those final sample (Table·2) included three to seven trials per in the other datasets considered in this study (0.62±0.21·m·s–1) individual, with a mean speed of 0.148±0.021·m·s–1 and a but using the same gait (fast walking trot; sensu Hildebrand, mean duty factor of 0.70±0.03. Gaits from steps used in the 1976). Data were collected from this alligator at 500·Hz as it force measurements were found to be essentially identical to walked down a runway and stepped with a single hind foot on those used in the treadmill data collection. To compare patterns a custom-built force platform (inserted so its surface was flush of GRF to other locomotor parameters, traces from typical with the trackway) that measured the three-dimensional GRF. GRF records were normalized to stance duration and aligned Using a mirror in the trackway, both lateral and dorsal views with kinematic, gait, and EMG data in Fig.·3 and with bone of the alligator were simultaneously filmed with a high speed strain data in Fig.·4. video camera (250·Hz, Kodak EktaPro model 1012) as it stepped on the platform. The positions of the limb joints were Femoral torsion digitized in the video frames and, combined with data on the Profiles of locomotor forces, kinematics and EMGs were magnitude and orientation of the GRF, the torsional moment compared to evaluations of femoral torsion based on in vivo of the GRF about the long axis of the femur was calculated bone strain recordings (Blob and Biewener, 1999). Bone using custom software. This moment indicates the tendency of strains were recorded by surgically implanting strain gauges on the GRF to rotate the femur. If the GRF is directed posterior the right femur of three subadult alligators (total length to the femoral long axis, it would tend to rotate the femur 0.98–1.04·m; body mass 1.73–2.27·kg) while the animals were medially or inward (counterclockwise if viewing the right under anesthesia (following protocols of Biewener, 1992). femur from its proximal end); if the GRF is directed anterior Rosette gauges were attached to the dorsal and ventral surfaces to the femoral long axis, it would tend to rotate the femur of the femur with cyanoacrylate adhesive, after the removal of laterally or outward (clockwise if viewing the right femur from a window of periosteum and cleaning of the attachment site its proximal end). To evaluate the timing and direction of with ether. Central elements of the rosette gauges were aligned torsional moments relative to limb kinematics, muscle activity, with the long axis of the femur and indicated longitudinal and bone strains, a representative trace of the torsional moment

THE JOURNAL OF EXPERIMENTAL BIOLOGY 1000 S. M. Reilly and others of the GRF was normalized for stride duration and aligned with stride duration) and ending just after mid-stance (38% of stride traces for other locomotor parameters (Fig.·3B). duration). Its onset coincides with the beginning of both hip adduction and knee extension in mid-swing phase (Fig.·3E). Thus, ILTIB1 contraction seems likely to contribute to Results protracting the thigh and extending the knee during swing Many features of the datasets we examined have been phase. However, ILTIB1 activity during stance phase described in previous studies (Gatesy, 1997; Reilly and Elias, coincides with a long period during which the knee and hip 1998; Blob and Biewener, 1999, 2001; Reilly and Blob, 2003; adduction angle are held nearly constant (Fig.·3E,F). Thus, Willey et al., 2004). For this analysis, our results will focus on ILTIB1 activity during stance phase appears to stabilize the hip the three major areas: (1) description of motor patterns of four and knee joints. hindlimb muscles for which detailed descriptions have not previously been published, (2) coordinated analysis of Puboischiofemoralis externus kinematic, gait, and motor pattern data during the swing and This ventral thigh muscle extends from the ventral surface stance phases of the locomotor cycle, and (3) the relative of the ischium to the proximal femur (Fig.·2C) and is in a timing and coordination of landmark features of kinematic, position to adduct the femur. Puboischiofemoralis externus 3 gait, muscle activity, GRF, and bone loading data from the (PIFE3) becomes active in the last quarter of swing phase and stance phase of the alligator hindlimb cycle. continues to fire for the first three-fourths of stance; thus, its activity coincides with the entire time during which the femur Hindlimb muscle activity patterns retracts. Because actual femoral adduction is slight during this Mean motor patterns for 12 muscles quantified in this study period, PIFE3 appears to serve as hip stabilizer during stance. and four from Gatesy (1997; indicated with asterisks) are presented in Table·1 and illustrated in Fig.·3F. For nearly all Gastrocnemius of the muscles that were examined by both Gatesy (1997) and Extending from the posterior aspect of the knee to the heel here (Table·1), motor patterns were remarkably similar (Fig.·2D) this muscle is clearly positioned to extend the ankle between the studies, with mean onsets and offsets differing by as well as flex the knee. It is active from just after foot down at most only 10% of stride duration. One exception was until almost the end (~90%) of stance phase. Ankle extension ILTIB2, for which we found burst durations to be longer by begins at about 35% of stance duration; thus, during the second almost 20% stride duration (offsets of 52.3% versus 33.5% of two-thirds of its activity period, the gastrocnemius (G) clearly stride duration). In addition, although some muscles recorded serves as an ankle extensor (plantarflexor). However, the ankle by Gatesy (1997) displayed pulsatile EMG signals, all of the flexes during the first third of the activity period of the muscles we recorded consistently showed discrete EMG bursts gastrocnemius. Contraction of the gastrocnemius during ankle rather than pulsatile signals. For FTE and FTI2 in particular, flexion probably serves to control the degree of ankle flexion we found discrete burst patterns that confirm the patterns and, thereby, stabilize the ankle prior to the onset of its identified by Gatesy (1997) from more sporadic bursts. Of the extension. 16 muscles under consideration, eight had a majority of their activity during stance phase (red in Fig.·2), six were active Tibialis anterior primarily during swing phase (blue in Fig.·2), and two were Extending from the proximal tibia to the metatarsals active in both phases (black in Fig.·2), the continuously active (Fig.·2D) Tibialis anterior (TA) is well-situated to act as an ILTIB1 and the PIT with discrete bursts in both phases. Reilly ankle flexor (dorsiflexor). Corresponding to this role, TA is and Blob (2003) provided the first summaries of motor patterns active in the first half of swing phase (68–91% of stride for four of these muscles (ILTIB1, PIFE3, G and TA) but their duration), essentially matching duration of ankle flexion during analysis focused on changes in burst characteristics related to swing (Fig.·3). However, the TA was not active during ankle limb posture, rather than propulsive dynamics. Therefore, we flexion in early stance phase suggesting that dorsiflexion at this will describe the motor patterns of these four muscles in detail time is a result of another muscle or passive effects of higher before outlining the new clarifications of muscle action limb movements. patterns derived from coordinated analyses of the 16 muscles considered in this study. Summaries of mean onset and offset Motor control of swing phase hindlimb movements times for each muscle are reported in Table·1 and illustrated in Six muscles were active primarily during swing phase Fig.·3F. (Fig.·2C,D: blue). Although no ground reaction forces are associated with this portion of the limb cycle as the leg Iliotibialis, head 1 protracts, the motor patterns of muscles active during this time Extending from the dorsal pelvis to the knee (Fig.·2B), can be compared to joint kinematics to evaluate muscular Iliotibialis, head 1 (ILTIB1) is in a position to abduct the femur function. and extend the knee. ILTIB1 has the earliest onset of any of the primarily stance phase muscles (Table·1), with activity Femoral abduction and knee flexion beginning just before the middle of swing phase (–18% of Of the many possible muscles in position to abduct the

THE JOURNAL OF EXPERIMENTAL BIOLOGY Alligator hindlimb function 1001

Fig.·3. Integration of five levels of Hindlimb stance dynamics analysis of stance phase dynamics A during hindlimb locomotion in Limb Support and Limb alligators. (A) Dynamic phases of loading propulsion unloading stance. (B) Torsional moment of the 0.1 B Medial (counterclockwise) ground reaction force acting on the 0 femur; illustrates torsional moments –0.1 Lateral (clockwise) Proximal view GRF on

on the right femur as viewed from the Torsional moment of acetabulum. (C) Three-dimensional femur (Nm) 0.6 ground reaction forces (top, vertical; C Vertical middle, fore–aft; bottom, 0.4 mediolateral) in body weight units (BWU). (D) Mean footfall patterns; R, 0.2 right; L, left; H, hindlimb; F, forelimb. Braking Propulsion (E) Mean hindlimb kinematics. (F) 0 forces (BWU)

Mean electromyographical patterns Ground reaction Medial for 16 hindlimb muscles, constructed –0.2 from the data in Table·1 and Gatesy RH t D i RF (1997; indicated by *). All of the data a

G LF (details in Materials and methods) are LH from alligators of similar size, moving 130 at the same speed (~0.146·m·s–1), 120 E Knee posture (femur adducted 51-60°), and 110 gait (duty factors 0.70–0.73). Vertical 100 n green bars delineate the three dynamic 90 Femoral retractio Ankle phases of limb function (from A) 80 during the stance phase: limb loading, 70 support and propulsion and limb 60 Adduction unloading, described in text. Limb joint angles (deg.) 50 Femoral adduction F 40 femur, only ILFIB and ILFEM F (Fig.·2C) were active primarily Swing Stance Swing during swing phase. Both muscles FTE FTE FTI2* initiate their activity in late stance FTI2* CFL* CFL* phase (Fig.·3F; first group of ILTIB1 ILTIB1 swing phase muscles). Activity in ILTIB2 ILTIB2 ILFEM, which is positioned to act FMTI FMTI solely as a femoral abductor, ends ADD1 ADD1 PIT PIT at about one third through the PIFE3 PIFE3 duration of swing phase, while G G activity in ILFIB, which passes ILFIB* ILFIB* over the knee joint, is active ILFEM ILFEM PIFI2 PIFI2 through nearly half of swing PIFE2* PIFE2* phase. These bursts coincide with AMB1 AMB1 the continuing abduction of the TA TA femur in early swing phase right –30 0 20 40 60 80 100 after the foot is raised from the Stride duration (%) ground (Fig.·3E). These bursts 0 25 50 75 100 also coincide with the swing phase Stance duration (%) burst of PIT, a femoral adductor. The synchronization of these bursts seems to indicate a Correspondingly, the period of ILFIB activity matches the controlled abduction (ILFEM) and adduction (PIT) of the rapid period of knee flexion during early swing phase femur early in swing phase before the femur is quickly (Fig.·3E). Therefore, ILFIB appears to contribute to both adducted back to its position at foot down by the stance phase femoral abduction and knee flexion in swing phase. bursts of PIT and other femoral adductors. With regard to ILFIB, Gatesy (1997) showed that with its low origin on the Femoral protraction and knee extension iliac blade and oblique crossing of the knee joint, ILFIB Two anteriorly positioned, proximal thigh muscles, PIFI2 is anatomically well positioned to flex the knee. and PIFE2, exhibited focused bursts of activity in the swing

THE JOURNAL OF EXPERIMENTAL BIOLOGY 1002 S. M. Reilly and others

Table·1. Patterns of hind limb muscle activity for alligators walking at 0.146·m·s–1 Muscle Abbreviation Onset (%) Offset (%) N Flexor tibialis externus FTE 0.9±3.0 52.6±4.5 2 Flexor tibialis internus 2* FTI2 –6.0±4.6 37.3±12.0 2* Caudofemoralis longus* CFL –6.0±0.0 55.9±0.0 4* Iliotibialis 1 ILTIB1 –18.0±6.5 38.0±7.3 2 Iliotibialis 2 ILTIB2 –10.9±7.2 52.3±5.3 2 Femorotibialis internus FMTI –8.4±1.9 48.2±3.2 3 Adductor femoris 1 ADD1 –9.4±4.1 38.7±5.5 3 Puboischiotibialis (stance burst) PIT –16.7±2.5 38.6±2.6 1 (swing burst) PIT 67.2±3.9 79.5±2.4 1 Puboischiofemoralis externus 3 PIFE3 –6.0±2.7 48.1±4.1 2 Gastrocnemius G 3.7±5.9 63.4±4.9 3 Iliofibularis* ILFIB 66.1±5.5 84.0±3.0 3* Iliofemoralis ILFEM 63.6±2.3 78.3±1.4 1 Puboischiofemoralis internus 2 PIFI2 67.6±6.5 92.7±3.4 1 Puboischiofemoralis externus 2* PIFE2 75.9±8.7 91.6±5.1 2* Ambiens 1 AMB1 78.0±3.8 97.3±1.8 2 Tibialis anterior TA 68.1±4.2 91.2±3.0 2

*Data for muscles studied by Gatesy (1997) moving at similar speeds (0.1–0.15·m·s–1) and posture (60° femoral adduction angle). Mean timing variables (±S.D.) are measured relative to the time of foot down for each stride and scaled to stride duration. Ten strides per individual averaging 51° of femoral adduction angle at mid-stance and 70% duty factor are included for each individual (N) per muscle. These patterns are illustrated relative to other aspects of limb dynamics in Fig.·3F. phase (Fig.·3F; second group of swing phase muscles). PIFI2 phase period of ankle flexion (Fig.·3E,F). Although G is is positioned to oppose the action of CFL (Fig.·2D); thus, PIFI2 anatomically located to act as a major ankle extensor, it is not probably serves to protract the femur and rotate it laterally (in active during swing phase ankle extension. However, AMB1 the opposite direction from CFL). Its activity closely parallels is active during mid to late swing phase, and its auxiliary the period of femoral protraction during swing phase. PIFE2 tendon to the heel appears to have an important role in is active in the latter half of femoral protraction during swing. extending the ankle prior to the placement of the foot on the Its ventral position (originating from the pubis) suggests it ground. could contribute to femoral adduction during swing phase, but its anterior position suggests that, like PIFI2, PIFE2 may act Motor control of stance phase hindlimb movements as a primary protractor of the femur. The ambiens (AMB1, Femoral retraction and rotation Fig.·2D) is also located in a position where its contraction Of the nine dorsally positioned thigh muscles considered in could protract the femur. It is active somewhat later than PIFI2 this analysis, only six were active during stance phase and PIFE2, during the period of knee extension that occurs (Fig.·2A,B). Of these, three (FTE, FTI2, CFL; Fig.·2A, Fig. 3F; during femoral protraction (Fig.·3E,F). Because AMB1 spans first set of three stance phase muscles) are positioned the knee as well as the hip (Fig.·2D), the timing of its activity posteriorly and represent a functional unit that could retract and indicates it serves as a primary swing phase knee extensor as rotate the limb. CFL has been identified as the primary femoral well as a limb protractor during late swing phase. Several other retractor and medial rotator during limb retraction in muscles that are anatomically positioned to extend the knee crocodilians (Gatesy, 1990, 1997). EMG data show that CFL also become active during mid to late swing phase as knee activity coincides with the entire period of femoral retraction, extension commences. ILTIB1 activity begins just after that of from its start during late swing phase until retraction stops AMB1 (Fig.·3F), indicating that ILTIB1 probably also nearly three quarters through the duration of stance phase contributes to the initiation of swing phase knee extension. (Fig.·3E,F). FTE and FTI2 are also located in positions suited ILTIB2 and FMTI become active toward the end of swing to contribute to femoral retraction, and were considered phase (Fig.·3F), so their contraction also may contribute to ancillary stance phase hip retractors by Gatesy (1997). With knee extension that occurs during this time (Fig.·3E). insertions on the lateral aspect of the upper shank (Fig.·2A), both could also contribute to femoral rotation and, potentially, Ankle flexion and extension knee flexion (see below). However, these muscles have The ankle undergoes first flexion, then extension during somewhat different motor patterns. FTI2 activity parallels the swing phase. TA is anatomically situated to flex the ankle, first two-thirds of the CFL burst, suggesting it contributes extending from the tibia to the dorsal aspect of the foot primarily to early retraction. Like CFL, FTI2 is situated to (Fig.·2D). It exhibits a strong EMG burst during the first two- contribute to medial femoral rotation; however, FTI2 is active thirds of swing phase, coinciding very closely with the swing primarily when the GRF would tend to rotate the femur

THE JOURNAL OF EXPERIMENTAL BIOLOGY Alligator hindlimb function 1003 laterally (i.e. in the opposite direction; Fig.·3B,F), suggesting (ADD1, PIT, PIFE3, Figs·2C, 3F; third set of three stance that this muscle might help to stabilize the femur against phase muscles) are anatomically situated to act as femoral rotation during early retraction. The same may be true for the adductors. These three muscles are active from mid to late CFL early in stance phase. In contrast, FTE has a later burst swing phase until well into stance phase, but their burst of activity, beginning at foot down and ceasing nearly patterns are somewhat staggered in time. PIT appears to act as simultaneously with the end of the CFL burst by the end of a swing phase femoral adductor, becoming active before the femoral retraction (Fig.·3E,F). The rotational moment of the middle of swing phase coincident with a major period of GRF shifts from lateral to medial during this time, acting in femoral adduction. There are two lines of evidence that the PIT the same direction as FTE and CFL by the midpoint of their actively powers adduction during the swing phase. First, the bursts. This suggests that FTE, like CFL, contributes to swing phase burst is associated with the complete repositioning femoral rotation as well as femoral retraction. of the limb in adduction during swing (Fig.·3E,F). Second, the swing phase PIT EMG intensity (mean area) is approximately Knee stabilization and knee flexion the same as that during stance phase (Reilly and Blob, 2003). FTI2 and FTE are anatomically situated to flex the knee joint In contrast, ADD1 and PIFE3 become active in the last third as well as retract the femur. However, during stance phase the to fourth of swing phase, after femoral adduction has stopped. knee joint is static before it flexes (Fig.·3E). Thus, FTI2, which All three muscles are active through most of the first three is active only while the knee is static, appears to stabilize the quarters of stance, but PIFE3 activity continues longer than knee rather than flex it. In contrast, FTE is active well into the that of the other muscles, lasting until femoral retraction stops period of knee flexion during the latter half of stance and, and vertical GRFs begin to decrease. Because the activity of therefore, probably helps to actively flex the knee. Although these muscles coincides with phases when the femur is static knee flexion continues into late stance (Fig.·3E), activity ceases (or actually abducted as much as 5°), these three muscles in both of these potential knee flexors well before the end of appear to stabilize the hip, counteracting the weight of the body the step (Fig.·3F). Body weight might passively contribute to as it is supported during stance phase. knee flexion during later portions of stance phase, but the hindlimb will tend to be unloaded when the next limb couplet Ankle extension of the trot contacts the ground later in stance phase (Fig.·3D; The remaining stance phase muscle studied was the G, see later Results). As mentioned above, knee flexion during which appears to be a primary ankle extensor (Fig.·1D; Fig.·3F; late stance might be actively controlled by the shank muscle last stance phase muscle). Our EMG data show that G activity gastrocnemius, which spans the flexor side of the knee joint begins after the start of stance and continues longer than that and is active until almost the end of stance (Fig.·3F). of any other stance phase muscle (until nearly 90% stance Three other dorsally positioned thigh muscles that are active duration). Its activity begins near the end of ankle flexion during stance phase (FMTI, ILTIB1 and ILTIB2; Figs·2B, 3F; during early stance and continues through the entire period of second set of three stance phase muscles) are aligned more ankle extension during stance phase. It is not active during closely with the long axis of the femur and share a wide swing phase; thus, another muscle must control ankle tendinous insertion extending to the proximal tibia. These extension that occurs while the foot is not in contact with the muscles all become active in mid to late swing phase as the ground. It is noteworthy that FTE not only has an auxiliary knee extends (Fig.·3E). However, both the onset and offset of tendon extending to the calcaneum, but that it has essentially ILTIB1 activity are shifted earlier relative to those of the other the same motor pattern as G. Thus, FTE seems likely to also two muscles by almost 20% stance duration. Unlike ILTIB1, contribute to stance phase ankle extension. both ILTIB2 and FMTI become active during the end of swing phase and fire through the first three quarters of stance phase. Hindlimb ground reaction forces Gatesy (1997) reported knee extension during the latter half of Typical three-dimensional GRF traces for alligator hindlimb stance phase and, therefore, described these three muscles as steps for animals moving the same speed as in the EMG studies stance phase knee extensors. However, because our results are presented in Fig.·3C. Means for key events of the force (and data on other speeds and postures; Reilly and Elias, 1998) patterns are presented in Table·2. Alligator hindlimb steps show the knee remaining static or flexing during stance phase, exhibited vertical force profiles that consistently increased to we conclude that these muscles act to stabilize the knee as it near peak values of about half of body weight in the first fifth accommodates and supports body weight through the first three of the stance phase. Near peak forces are maintained until quarters of stance. In fact, it is not until activity ceases in these about the last third of stance phase when body support wanes. muscles that stance phase knee flexion becomes substantial, Fore–aft forces indicate a brief braking component in the first further suggesting that these muscles act to limit knee flexion fifth (15.4%) of stance phase with maximum braking force of during stance and that late in stance they no longer counter the 5.4% of body weight at 6% of stance duration. Thereafter, a effects of body weight and flexor muscles on the knee joint. long gradually increasing propulsive component occurs during the middle (from 15.4 to 64.4%) of stance phase coincident Femoral adduction with the period when vertical forces plateau. Maximum The last three thigh muscles active during stance phase propulsive force of 5.9% of body weight occurs at 64.4% of

THE JOURNAL OF EXPERIMENTAL BIOLOGY 1004 S. M. Reilly and others stance duration. Propulsive effort then decreases to around zero (coincident with the left vertical shaded bar in Figs·3, 4). Joint early in the last third of stance duration. Mediolateral GRFs kinematics (Fig.·3E) show that the knee and ankle both flex as are consistently negative (i.e. medially directed), and these the limb is loaded. However, as the femur is retracted during forces reach maximum values of about 10.6% of body weight this phase (continuing motion that began during the previous in the first fifth (19.1%) of stance duration and then gradually decrease over the rest of the stance phase, dropping to zero just Hindlimb stance phase before the foot is picked up. Limb Support and Limb Hindlimb force vectors loading propulsion unloading Ground reaction forces were reconfigured to reflect hindlimb A output forces. Sagittal (Fig.·4A) and transverse (Fig.·4B) two- 100 dimensional limb force vectors illustrate how the hindlimb pushes on the ground during the stance phase. Once the foot has settled in after impact the net hindlimb vertical force vector 90 is directed about 10° anterior to vertical (~80° on Fig.·4A) and Sagittal about 60° lateral to the direction of travel (Fig.·4B). In the first Vertical/foreaft 80 one-fifth of stance phase the hindlimb force vector swings posteriorly to about 4° posterior to vertical (~94° in Fig.·4A) and 110° relative to the direction of travel. Throughout the 90° 70 middle portion of stance, the sagittal force vector then remains B relatively constant (about 95°) whereas the transverse 130 component swings continually more posteriorly (from 110–130°) until the greatest posterolateral force direction 110 occurs at about two-thirds of stance phase. During the last quarter of stance phase, the longitudinal propulsive component 90 Orientation of hindlimb force vector (deg.) becomes negligible (Fig.·3C) and the sagittal hindlimb force Transverse Lateral/foreaft vector swings anteriorly to about 90° (Fig.·4A). At the same 70 time, the transverse forces are dominated by a lateral push on 90° the ground by the hindlimb. This indicates that the last portion 50 0255075 100 of the stance phase (after the time that the fore–aft forces return Stance phase (%) to zero; Fig.·3C) involves only vertical and lateral forces. 300 Because of the relatively small mediolateral forces, the limb is C 0 primarily supporting the body with a secondary role of providing mediolateral stability. The final noisy shifts in –300 hindlimb force orientation just prior to lift-off are –600 inconsequential as they are associated with increasingly trivial ) 900 force magnitudes. µε D 600 Integrative dynamics of stance phase for the alligator 300 hindlimb Our coordinated analyses of hindlimb kinematic, gait, motor 0

Femoral strain ( –100 pattern, force and bone loading data during stance in alligators 300 (Figs·3, 4) indicate three distinct dynamic phases during this E portion of the locomotor cycle for the hindlimb. Axial0 Shear Principal

Limb-loading phase Fig.·4. Typical two-dimensional hindlimb force vectors and femoral During the first fifth of the step, the hindlimb is loaded strains for alligators using a walking trot. Orientations of the (A) as body weight is transferred fully to the stance phase sagittal and (B) transverse hindlimb force vectors are shown in forelimb–hindlimb couplet. This limb-loading phase is relation to the dynamic hindlimb stance phases shown in Fig.·3 (vertical bars). Note that these vectors are equal in magnitude but distinguished by several dynamic features. First, loads on the opposite in direction to the components of the ground reaction force; femur (axial, principal, and shear strains, Fig.·4C–E) increase thus, when the hindlimb force vector is directed posteriorly (>90°), throughout this phase, rising toward maxima nearly the ground reaction force is directed anteriorly. (C–E) In vivo simultaneously by the end of this portion of the step. Gait data principal, shear, and axial strains recorded from the alligator femur (Fig.·3D) show that this phase begins when the hindlimb during the stance phase (at 0.37 m·s–1; from Blob and Biewener, contacts the substrate and continues until the limbs of the 1999). (C,D) Strain traces from the dorsal femur. (E) Strain trace from opposing forelimb–hindlimb couplet begin their swing phases the anterior femur.

THE JOURNAL OF EXPERIMENTAL BIOLOGY Alligator hindlimb function 1005 swing phase), femoral adduction remains fairly constant. All Table·2. Timing and magnitudes of hindlimb force patterns of the muscles that will be active during the following for high walking alligators propulsive phase have started to contract by the end of the Variable Mean ± S.D. limb-loading phase; in fact, many of these muscles (including CFL, the major limb retractor) start to contract at the end of Maximum vertical force 0.532±0.027 the swing phase prior to limb loading (Fig.·3F). During the Time to maximum vertical force 35.8±2.4 Maximum braking force 0.054±0.018 limb loading period, the vertical component of the GRF rises Time to maximum braking force 6.0±0.7 to near maximum levels (>40% body weight), the medial Time to braking-to-propulsive transition 15.4±0.9 component of the GRF rises to its maximum (10.6% body Maximum propulsive force* –0.059±0.125 weight at 19% stance phase), and a small hindlimb braking Time to maximum propulsive force 64.4±5.4 impulse occurs (Fig.·3C, Table·2). When force components are Maximum lateral force** –0.106±0.011 recalculated to reflect the orientations of forces that the Time to maximum lateral force 19.1±1.4 hindlimb exerts on the ground (Fig.·4A,B), sagittally oriented hindlimb forces reflect deceleration during most of the limb- *Negative fore–aft values are accelerative forces. loading phase (i.e. the foot pushing anteriorly on the ground, **Negative mediolateral values are lateral forces. Fig.·4A), and transversely-oriented hindlimb forces transition Forces are in body weight units and times are in percentage of from anterolateral at the start of the phase to posterolateral by stance duration. Data are from 20 three-dimensional hindlimb force records pooled the end of the phase (Fig.·4B). In addition, the torsional across four individuals (N per individual=3, 5, 5, 7) averaging 0.148 moment of the GRF rises towards its maximum in the lateral m·s–1 (range 0.110–0.167·m s–1) to match the speed of kinematic and direction during the limb-loading phase (clockwise if the right gait data (average 0.146 m·s–1) as closely as possible. femur is viewed from its proximal end; Fig.·3B), opposite of the direction of rotation that would be expected to result from CFL contraction. posteriorly oriented; Fig.·4B). Limb joint kinematics show that Support-and-propulsion phase the femur continues to retract during this phase, but that the The middle portion of stance phase (between the vertical knee, ankle and femoral adduction angles change little during shaded bars in Figs·3 and 4) is distinguished by several features the first two-thirds of support and propulsion (Fig.·3E). In the indicating that hindlimb contributions to body support and last third of the support-and-propulsion phase, femoral propulsion are maximized during this phase. High vertical retraction continues at the same rate while the knee begins to forces are maintained throughout this phase (Fig.·3C), flex and the ankle begins to extend (plantarflex); thus, indicating more or less constant support of body weight by the shortening of the functional length of the limb caused by knee hindlimb. However, bone strains decrease throughout this flexion is counteracted by ankle extension. Consequently, phase (Fig.·4C–E) despite the fact that gait diagrams show that femoral retraction (which stops at the end of this phase) is the body is supported only by a single limb couplet during this primarily responsible for overall changes in hindlimb position entire phase (Fig.·3D). For the first half of the support and during the support-and-propulsion phase in alligators. During propulsion phase, while the GRF is still increasing (Fig.·3C), support-and-propulsion phase, the hip and knee move decreases in strain probably reflect an increasingly closer anteriorly relative to the foot. As a result, when the femur is alignment between the GRF and the femur. During the second nearly perpendicular to the body of the alligator (95-100° half of the support and propulsion phase, the orientations of femoral retraction angle, Fig.·3E), the GRF will shift from the GRF and femur will tend to diverge, but strains continue being directed anterior to the long axis of the femur (as in the to decline in correspondence with decreases in GRF limb-loading phase) to being directed posterior to the long axis magnitude. of the femur (Blob and Biewener, 2001). As this shift occurs Virtually all of the propulsive impulse occurs during this by the midpoint of the support-and-propulsion phase (close to phase (Fig.·3C) with increases to near maximal levels early in the time of peak vertical force; Fig.·3B), the torsional moment this phase. The medially directed GRF decreases from its of the GRF about the femur shifts from exerting a lateral maximum (of about 6% of body weight) at the onset of this moment to a medial moment (counterclockwise when the right phase to about half maximal values by the end of the support- femur is viewed from its proximal end; Fig.·3B). This moment and-propulsion phase (~64.4% of stance). Ratios of vertical reaches its maximum by the end of the support-and-propulsion and lateral hindlimb forces to fore-aft hindlimb forces show phase, and it acts to produce femoral rotation in the same that the foot exerts a more or less constant but slightly direction that is expected from contraction of the CFL. Thus, posteriorly directed force on the ground during this phase the actions of both the GRF and CFL have the potential to (Fig.·4A). However, throughout this phase, increases in the induce substantial medial rotation of the femur during its rearward force exerted by the foot, and coincident decreases in retraction in the support-and-propulsion phase. It should be the lateral force it exerts, result in a 20° posteromedial shift in noted, however, that shear strain magnitudes have declined the direction of the force that the hindfoot exerts on the ground from their maxima by the time during the step that both CFL (i.e. transverse forces applied by the hind foot become more and the GRF would induce femoral rotation in the same

THE JOURNAL OF EXPERIMENTAL BIOLOGY 1006 S. M. Reilly and others direction (Fig.·4D). Moreover, CFL activity ceases by the end limb-loading phase (Figs·3C, 4A,B) relate primarily to femoral of this phase (Fig.·3F). retraction and rotation (see also Gatesy, 1991; Blob and Biewener, 2001). Limb-unloading phase In the alligator femur, principal, shear and longitudinal Dynamic changes in the last third of stance phase delineate strains all reach their highest values early in the step by the end the unloading of the hindlimb at the end of the step as the of the limb-loading phase or the beginning of the support-and- opposite forelimb-hindlimb couplet touchdown and begin their propulsion phase (Fig.·4C–E). These peak strains coincide with loading phase (Fig.·3D). Limb bone strains decline to zero several events, including the point when body weight becomes (Fig.·4C–E) as does propulsive force. The mediolateral fully supported by a single limb couplet (Fig.·3D), the onset component of the GRF decreases to zero during the second half of the plateau in vertical hindlimb forces (Fig.·3C), the of this phase. Consequently, during the first half of the development of peak medial GRFs acting on the hindlimb unloading phase, the sagittal limb forces become more purely (Fig.·3C), the end of the hindlimb braking impulse (Fig.·3C), vertically oriented as the transverse limb forces become more and the peak lateral (outward) rotational moment of the GRF lateral (Fig.·4A,B). The final, more irregular, portion of the (Fig.·3B). Thus, peak bone loads (i.e., maximum strains) are unloading phase reflects rolling over the hindfoot toes. Ratios associated with peak mediolateral forces and moments and the of vertical and lateral hindlimb forces to fore-aft hindlimb initial loading of body weight onto the limb just prior to the forces decrease from maxima (respectively 130° and 98°, support-and-propulsion phase, rather than with the highest Fig.·4A,B) to minima (~90°) in the first half of this phase, forces associated with propulsion or the support of body reflecting the decline of all force components to zero toward weight. Strain magnitudes actually decrease rapidly during the the end of the step. Most muscles active during the support- support-and propulsion-phase, even though the body is and-propulsion phase cease activity very early in the limb completely supported by one limb couplet and all hindlimb unloading phase, with the exception of the knee flexor/ankle stance phase muscles are active. Principal and shear strains extensor gastrocnemius, which is active for nearly the first ultimately return to zero by the end of the support-and- three quarters of this phase (Fig.·3F). Joint kinematics show propulsion phase (Fig.·4C,D) as the opposite limb couplet that the femur is held in a maximally retracted position during contacts the ground and begins to support body weight the limb-unloading phase, but that knee flexion and ankle (Fig.·3D). Axial strain, however, decreases to zero later during extension continue to counteract each other throughout this the step than shear strains. This suggests a decline in the phase (Fig.·3E), so that as vertical forces decline the functional importance of torsion relative to bending and axial loads later length of the limb distal to the knee changes very little in in the step (perhaps as the leg becomes more closely aligned length. The rotational moment about the long axis of the femur with the GRF), although this occurs at strain magnitudes much also falls to zero by the end of this phase (Fig.·3B). lower than the peak loads experienced earlier in the step.

Mechanisms underlying torsional loading of the alligator Discussion femur A major motivation for the synthesis of several aspects of Alignment of rotational moments of the GRF with shear alligator limb function in this study was to evaluate how strains and muscle activity patterns provides insight into the torsional loading of the femur was produced in this species. mechanisms underlying torsional loading of the femur in However, the integration of these datasets also allows us to alligators. Strain records directly indicate the actual refine evaluations of how several limb muscles contribute to deformation of the bone surface, and thus reflect the net loads body weight support and propulsion in alligators, and it resulting from all forces acting on the bone. Shear strains provides insights into the terrestrial locomotor mechanics of (reflecting torsion) increase to their maxima by the end of the vertebrates that do not use strictly parasagittal limb posture. limb-loading phase of stance (Fig.·4D) and indicate a medial twisting of the distal end of the femur (Blob and Biewener, Propulsive forces, in vivo strains, and the hindlimb loading 1999). Such twisting could be induced either by the contraction phase of limb retractor muscles with ventral insertions on the femur, At touchdown, when an alligator first places its hindfoot on by the rotational moment of the GRF if the GRF is directed the ground, some of its limb joints begin to flex, compressing posterior to the long axis of the femur or, perhaps, by combined the functional length of the limb as it begins to support the action of both processes. Femoral retractors such as CFL, FTI2 weight of the body. Although the knee flexes briefly just after and FTE are active at the end of the limb-loading phase when foot down, most of the shortening of functional hindlimb shear strains are at their highest levels (Fig.·3F), and thus these length during the limb-loading phase results from ankle muscles appear to contribute to the generation of torsional flexion. By the end of the limb-loading phase, every stance loads on the femur with a tendency to rotate the femur phase muscle examined was fully active, and the activity of medially. However, when shear strains are highest the GRF is these muscles held the ankle, knee and hip adduction angles directed anterior to the long axis of the femur (Blob and nearly constant while the femur is retracted and rotated. Thus, Biewener, 2001) and the GRF exerts a moment that would tend the limb motions that elicit propulsive GRFs by the end of the to rotate the femur laterally (Fig.·3B), i.e. in the opposite

THE JOURNAL OF EXPERIMENTAL BIOLOGY Alligator hindlimb function 1007 direction from rotation induced by the limb retractor muscles. Alligators differ in that medial rotation of the femur occurs This means that the unusually high shear strains seen in the with little change in knee angle. alligator femur must be produced by contraction of CFL and Generalized tetrapod hindlimb kinematics also appear to be other muscles against the rotational moment of the GRF. This controlled by muscle activity patterns that differ from those conclusion differs from the hypothesis of Blob and Biewener used by alligators. For example, the sprawling lizard (2001), who had suggested that high torsional loads in alligator Sceloporus clarkii exhibits distinct EMG patterns in which (1) limb bones result because both the GRF and CFL would act onset of G activity begins late in swing phase to actively extend additively and induce a moment that would tend to rotate the the ankle before the foot is placed on the ground, (2) FMTI has femur in the same direction. The GRF and CFL do act a swing phase burst to extend the knee before placing the foot additively to produce medial femoral rotation later in stance on the ground, and (3) TA has a large burst during early stance but only after shear strains have begun to decline from their that could help to pull the body over the foot (Reilly, 1995). peak. In fact, by the time the GRF exerts its maximum None of these EMG patterns occur in alligators (Fig.·3F). rotational moment in the medial direction at the end of the Even the common muscular function of using the CFL to support-and-propulsion phase (Fig.·3B), activity in the limb power hindlimb retraction in alligators and more sprawling retractor muscles ceases (Fig.·3F). Thus, additive action of the taxa (Peters and Goslow, 1983; Reilly, 1995; Gatesy, 1997) GRF and limb retractors (such as CFL) is not a primary factor could have a different effect on femoral loading because of in the generation of high torsional loads in the alligator femur differences in limb positions used by these animals. In highly near the end of the limb-loading phase. sprawling taxa that place the foot far forward at the beginning of stance, the GRF may be directed posterior to the long axis Is femoral torsion a general feature of non-parasagittal of the femur throughout the duration of stance. If true, then the locomotion? rotational moment of the GRF and CFL would be in the same The predominance of torsion as a mode of limb bone loading direction (both medial). Such an orientation of the GRF might is an unusual feature that distinguishes alligators and iguanas generate low torsional loads (reduced femoral shear strains) among highly sprawling species that initially place the foot far from most tetrapods in which limb bone loads have been anteriorly, as was found in alligators during the support-and- evaluated during terrestrial locomotion (Blob and Biewener, propulsion phase. Thus, high torsional loads might be a derived 1999). Blob and Biewener (1999) suggested that, if limb bone feature found among some non-parasagittal taxa including torsion was a common feature of limb bone loading among alligators, as opposed to a general feature of non-parasagittal crocodilians and lepidosaurs (the broader clades to which locomotion. Rather than reflecting an ancestral condition, the alligators and iguanas belong), then it might be a general and, limb kinematics, muscle activity patterns, and femoral loading perhaps, ancestral feature of terrestrial locomotor mechanics patterns of alligators appear to be derived relative to those of among species that do not use strictly parasagittal limb other non-parasagittal tetrapods. movements. However, alligators (and iguanas) exhibit several Two features of the locomotor apparatus of alligators may distinctive kinematic features that are not seen among taxa that result in reductions in overall locomotor effectiveness. First, habitually use either more or less sprawling limb posture. For alligators limit knee flexion-extension and femoral adduction example, salamanders, sprawling lizards and mammals all during stance phase. Instead, hindlimb retraction during the share a distinct pattern of knee kinematics in which flexion is support-and-propulsion and limb unloading phases is followed by extension during stance phase (Peters and Goslow, associated primarily with femoral medial rotation (together 1983; Ashley-Ross, 1995; Reilly, 1995, 2000), whereas with ankle plantarflexion). Consequently, effective vaulting alligator and iguana knees are held fixed in early stance and over the stance limbs may be limited. Indeed, walking then either extend or flex (Fig.·3; Brinkman, 1980; Reilly and efficiency is reduced in alligators: although the center of mass Elias, 1998; Blob and Biewener, 1999). In addition, at the rises and falls with each step, little external mechanical energy beginning of stance, hip protraction, knee extension, and ankle is recovered by the pendulum-like exchange of kinetic and extension are all greater (by 7–35°, 35–55°, and 30–45°, gravitation potential energies (19.8±2.0%; Willey et al., 2004). respectively) in salamanders and sprawling lizards (Ashley- The second unusual feature of alligator terrestrial locomotion Ross, 1994b; Reilly and Delancey, 1997b; Irschick and Jayne, is that the tail is dragged behind the body rather than elevated 1999) than in alligators and iguanas. Thus, whereas alligators off of the ground. The long, heavy tail causes the center of and iguanas place the hindfoot under or behind the knee at mass of alligators to lie more caudally (just in front of the touchdown, more sprawling taxa (e.g. salamanders and pelvis), so that body weight support is concentrated over the sprawling lizards) use greater limb joint extension at hindlimb (52%) during locomotion (Willey et al., 2004). The touchdown and are hypothesized to flex these joints to actively constant braking impulse of the tail is countered by propulsive pull the body over the foot (Ashley-Ross, 1994b, 1995; Reilly efforts of the limbs. Indeed, in order to balance the braking and Delancey, 1997b). Such comparisons of tetrapod impulse of the tail, the hindlimb exerts four times the kinematics led Ashley-Ross (1994b) to conclude that, in accelerative impulse needed to balance the braking impulse of conjunction with femoral retraction, knee flexion and extension the forelimb alone (Willey et al., 2004). These necessary are plesiomorphic features of the tetrapod hindlimb cycle. modifications in hindlimb function may further limit the ability

THE JOURNAL OF EXPERIMENTAL BIOLOGY 1008 S. M. Reilly and others of alligators to effectively use vaulting mechanics to recover supplying the alligators and the Ohio University Lab Animal external mechanical energy Resources for their outstanding success in maintaining the It is noteworthy that iguanas, which also exhibit substantial alligator colony. E. Anderson, A. Back, B. Brad, A. Clifford, femoral rotation (and torsion; Blob and Biewener, 1999) also D. Croft, J. Elias, R. Essner, K. Hickey, J. Keith, J. Kohler, L. drag large tails behind the body. In contrast, sprawling lizards Krautter, H. Larsson, P. Magwene, J. Noor, A. Parchman and that lift the tail during locomotion show net propulsive J. Tat assisted in data collection. Special thanks to J. Bertram, impulses of the hindlimb that balance the net braking impulses A. Biewener, M. Carrano, W. Corning, N. Espinoza, J. of the forelimb (our unpublished data) and much (two to three- Hopson, M. LaBarbera, J. Socha, M. Temaner and the Ohio fold) greater mechanical efficiency than alligators when University Evolutionary Morphology group, especially K. walking (Farley and Ko, 1997; S. Reilly, K. Hickey and A. Earls, A. Lammers, N. Stevens, E. Thompson, L. Witmer, for Parchman, unpublished data). With the tail raised, the their advice and help in many ways. This work was supported hindlimbs are only required to counteract the decelerative by an Ohio University Research Challenge grant, two Ohio impulse of the limbs and, thus, lizards may exhibit less axial University Honors Tutorial College Summer Research rotation of the femur and, thereby, lower torsional loads. Fellowships, the Clemson University Department of Experimental verification of these hypotheses remains to be Biological Sciences, and National Science Foundation grants performed. However, the results of our analyses suggest the IBN 9520719 (R.W.B.), IBN 9723768 (A.R.B.), and IBN possibility that patterns of femoral loading in quadrupedal, 9727212 and IBN 0080158 (S.M.R. and A.R.B.). terrestrial tetrapods may have diversified through evolutionary changes in several lineages, rather than along a single path. One consequence of the advent of limb rotation in alligators References is that several muscles that actively shorten to flex and extend Alexander, R. McN. (1974). The mechanics of a dog jumping, Canis familiaris. J. Zool. Lond. 173, 549-573. limb joints during stance phase in sprawling (salamanders; Alexander, R. McN. and Jayes, A. S. (1983). A dynamic similarity Ashley-Ross, 1994b; lizards: Reilly, 1995; Reilly and hypothesis for the gaits of quadrupedal mammals. J. Zool. Lond. 201, 135- DeLancey, 1997b) and erect quadrupeds (mammals; Goslow 152. Ashley-Ross, M. A. (1994a). Metamorphic and speed effects on hindlimb et al., 1973, 1981; Halbertsma, 1983; Smith et al., 1993; kinematics during terrestrial locomotion in the salamander Dicamptodon Vilensky and Gankiewicz, 1990) must now act in isometric or tenebrosus. J. Exp. Biol. 193, 285-305. even eccentric contraction to stabilize the knee and ankle Ashley-Ross, M. A. (1994b) Hindlimb kinematics during terrestrial locomotion in a salamander (Dicamptodon tenebrosus). J. Exp. Biol. 193, during the support-and-propulsion phase. Although hindlimb 255-283. EMG data are lacking for iguana, the knee and ankle flexors Ashley-Ross, M. A. (1995). Patterns of hindlimb motor output during walking in alligators are clearly modulated to stiffen the limb during in the salamander Dicamptodon tenebrosus, with comparisons to other tetrapods. J. Comp. Physiol. A. 177, 273-285. limb rotation. In addition, the same basic leg stiffening pattern Bakker, R. T. (1971). Dinosaur physiology and the origin of mammals. is modulated to maintain different fixed joint angles during Evolution 25, 636-658. stance phase across a range of postural heights (Reilly and Biewener, A. A. (1989). Scaling body support in mammals: limb posture and muscle mechanics. Science 245, 45-48. Blob, 2003). This adds to other recent discoveries (Gillis and Biewener, A. A. (1990). Biomechanics of mammalian terrestrial locomotion. Biewener, 2001, 2002, 2003) that counter common Science 250, 1097-1103. assumptions that homologous muscles are used at the same Biewener, A. A. (1992). In vivo measurement of bone strain and tendon force. In, Biomechanics – Structures and Systems: A Practical Approach (ed. A.A. time in the limb cycle across quadrupeds, or to move joints in Biewener), pp. 123-147. New York: Oxford University Press. the same way. In addition, motor patterns in alligator reveal Biewener, A. A. and Dial, K. P. (1995). In vivo strain in the humerus of the presence of local and temporal segregation of muscle pigeons (Columba livia) during flight. J. Morphol. 225, 61-75. Biewener, A. A., Thomason, J., Goodship, A. and Lanyon, L. E. (1983). functions during locomotion. Local segregation is evident in Bone stress in the horse forelimb during locomotion at different gaits: a muscles lying side by side (such as the ILTIB1, ILTIB2 and comparison of two experimental methods. J. Biomech. 16, 656-576. AMB1) that are dedicated to performing different functions in Biewener, A. A., Thomason, J. and Lanyon, L. E. (1988). Mechanics of locomotion and jumping in the horse (Equus): in vivo stress in the tibia and the stance (knee stabilization in the former two) and swing metatarsus. J. Zool. Lond. 214, 547-565. phases (knee extension in AMB1) of the stride. Furthermore, Blickhan, R. and Full, R. J. (1992). Mechanical work in terrestrial only two (PIT, ILTIB1) of our 16 muscles had clear functions locomotion. In Biomechanics: Structures and Systems (ed. A. A. Biewener), pp. 75-96. New York: Oxford University Press. in both stance and swing phases. Most notably, PIT appears to Blob, R. W. (2001). Evolution of hindlimb posture in nonmammalian aid other muscles (ADD1, PIFE3) in supporting the body therapsids: biomechanical tests of paleontological hypotheses. Paleobiol. weight during the stance phase, while it is alone in 27, 14-38. Blob, R. W. and Biewener, A. A. (1999). In vivo locomotor strain in the counteracting femoral abduction (by the ILFIB, ILFEM) hindlimb of Alligator mississippiensis and Iguana iguana: implications for during swing phase. These results point to several future the evolution of limb bone safety factor and non-sprawling limb posture. J. directions for research on iguanas, sprawling lizards and semi- Exp. Biol. 202, 1023-1046. Blob, R. W. and Biewener, A. A. (2001). Mechanics of limb bone loading erect mammals that could improve understanding of the effects during terrestrial locomotion in the green iguana (Iguana iguana) and of posture, tail dragging, and phylogeny on the diversity of American alligator (Alligator mississippiensis). J. Exp. Biol. 204, 1099- quadrupedal locomotor mechanisms. 1122. Brinkman, D. (1980). The hindlimb step cycle of Caiman sclerops and the mechanics of the crocodile tarsus and metatarsus. Can. J. Zool. 58, 2187- We thank Ruth Elsey at the Rockefeller Wildlife Refuge for 2200.

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